RESUMEN
The physiological functions of the cellular prion protein, PrP(C), as a cell surface pleiotropic receptor are under debate. We report that PrP(C) interacts with vitronectin but not with fibronectin or collagen. The binding sites mediating this PrP(C)-vitronectin interaction were mapped to residues 105-119 of PrP(C) and the residues 307-320 of vitronectin. The two proteins were co-localized in embryonic dorsal root ganglia from wild-type mice. Vitronectin addition to cultured dorsal root ganglia induced axonal growth, which could be mimicked by vitronectin peptide 307-320 and abrogated by anti-PrP(C) antibodies. Full-length vitronectin, but not the vitronectin peptide 307-320, induced axonal growth of dorsal root neurons from two strains of PrP(C)-null mice. Functional assays demonstrated that relative to wild-type cells, PrP(C)-null dorsal root neurons were more responsive to the Arg-Gly-Asp peptide (an integrin-binding site), and exhibited greater alphavbeta3 activity. Our findings indicate that PrP(C) plays an important role in axonal growth, and this function may be rescued in PrP(C)-knockout animals by integrin compensatory mechanisms.
Asunto(s)
Axones/metabolismo , Integrinas/metabolismo , Proteínas PrPC/metabolismo , Vitronectina/metabolismo , Animales , Sitios de Unión , Línea Celular , Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Ganglios Espinales/citología , Ganglios Espinales/embriología , Humanos , Ratones , Proteínas PrPC/química , Unión Proteica , Mapeo de Interacción de Proteínas , Estructura Terciaria de Proteína , Vitronectina/químicaRESUMEN
In order to predict the binding regions within the complex formed by Toll-like receptor 5 (TLR-5) and flagellin, a complementary hydropathy between the two proteins was sought. A region common to the flagellins of Salmonella enterica serovar Typhimurium, Pseudomonas aeruginosa, and Listeria monocytogenes was shown to be hydropathically complementary to the 552-to-561 fragment of TLR-5, whose sequence is EILDISRNQL. The hydrophobicity profile of this region is shared with flagellins of 377 bacterial species out of a total of 723 publicly available sequences. A conformational analysis of the predicted binding site of TLR-5, whose structure is still unknown, was carried out with a methodology already applied to similar problems. To sample the conformations available to the peptide chain, a plot of the number of conformations per unit energy interval (density of states) versus energy was built. Following a theoretical argument, conformations belonging to maxima in this plot were selected. The most stable structure obtained in this search, an alpha-helical conformation, was shown to form the electrostatic interactions Glu552-Gln89, Asp555-Arg92, and Arg558-Glu93 with the predicted binding site of the flagellin of S. enterica serovar Typhimurium, formed by the 88-to-97 chain fragment (LQRVRELAVQ), which is likewise alpha helical.
Asunto(s)
Flagelina/química , Flagelina/metabolismo , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/metabolismo , Receptores de Superficie Celular/química , Receptores de Superficie Celular/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Flagelina/genética , Interacciones Hidrofóbicas e Hidrofílicas , Listeria monocytogenes/química , Listeria monocytogenes/genética , Listeria monocytogenes/metabolismo , Glicoproteínas de Membrana/genética , Modelos Moleculares , Datos de Secuencia Molecular , Unión Proteica , Conformación Proteica , Pseudomonas aeruginosa/química , Pseudomonas aeruginosa/genética , Pseudomonas aeruginosa/metabolismo , Receptores de Superficie Celular/genética , Salmonella typhimurium/química , Salmonella typhimurium/genética , Salmonella typhimurium/metabolismo , Programas Informáticos , Receptor Toll-Like 5 , Receptores Toll-LikeRESUMEN
Various procedures are employed to relate the structural tendencies of polypeptide chain fragments to amino acid residues that in general have low background frequencies. A numerical evaluation of the content of these amino acids, named amino acid diversity, is defined. Distributions of the amino acid diversity parameter in databases containing exons, introns, and randomized exons show that there is a small difference between exons and shuffled exons, a detectable difference between exons and introns, and a large difference between exons and totally randomized exons.